The COVID-19 pandemic caused by SARS-CoV-2 represents a public health emergency, which became even more challenging since the detection of highly transmissible variants and strategies against COVID-19 were indistinctly established. We characterized the temporal viral load kinetics in individuals infected by original and variant strains. Naso-oropharyngeal swabs from 33,000 individuals (admitted to the IRCCS Santa Lucia Foundation Drive-in, healthcare professionals and hospitalized patients who underwent routinary screening) from November 2020 to June 2021 were analyzed. Of them, 1735 subjects were selected and grouped according to the viral strain. Diagnostic analyses were performed by CE-IVD RT-PCR-based kits. The subgenomic-RNA component was assessed in 36 subjects using digital PCR. Infection duration, viral load decay speed, effects of age and sex were assessed and compared by extensive statistical analyses. Overall, infection duration and viral load differed between the groups (p < 0.05). Male sex was more present among both original and variant carriers affected with high viral load and showing fast decay speed, whereas original strain carriers with slow decay speed resulted in older (p < 0.05). Subgenomic-RNA was detected in the positive samples, including those with low viral load. This study provides a picture of the viral load kinetics, identifying individuals with similar patterns and showing differential effects of age and sex, thus providing potentially useful information for personalized management of infected subjects.